Invasive Germ Cell Tumors Research Articles

Bilateral Gonadoblastoma in a 13-Year-Old Female with Turner Syndrome and Y Chromosome: A Case Report

Abstract Introduction/Objective Gonadoblastoma is a relatively rare type of ovarian tumor that has a high risk for patients with 45X/46XY. It is derived from the Y chromosome at its long arm GBY region, which contains the most likely candidate gene called testis-specific protein Y-encoded (TSPY). Methods/Case Report In this case report, we present a case of a 13-year-old girl with Turner syndrome whose cytogenetic test showed 45X with monosomy X and a siodicentric Yq chromosome. As she carried a high risk for the development of gonadoblastoma due to the presence of the Y chromosome and clinically had premature ovarian failure, she received further imaging studies, which did not reveal obvious ovarian masses. She underwent resections of bilateral fallopian tubes and ovaries. Bilateral fallopian tubes were unremarkable. However, within the bilateral ovarian tissue, foci of gonadoblastoma were found (0.5 to 0.6 cm), composed of large round germ cells (confirmed by positive OCT 3/4 and SALL4 stains) intermingled with oval-shaped sex cord-stromal cells forming tubular/cribriform structures (confirmed by positive staining for inhibin, calretinin, and FOXL2). The stromal cells of gonadoblastoma, but not germ cells, were positive for androgen receptor staining, while both components were negative for estrogen receptor staining. No invasive germ cell tumor or sex cord tumor was present. Results (if a Case Study enter NA) N/A Conclusion The positive androgen receptor staining in stromal cells of the gonadoblastoma supports the view that the Y chromosome-driven androgen system may play a role in the development of gonadoblastoma in a bi-cellular pattern.

Prognostic Value of Apoptosis-Inducing Factor (AIF) in Germ Cell Tumors.

Simple SummaryGerm cell tumors (GCTs) are the most common solid malignancies in young men. GCTs are extraordinary sensitive to chemotherapy and represent a model of curable cancer. However, in a small proportion of patients the disease progresses or relapses despite administration of salvage chemotherapy. Apoptosis is a form of programmed cell death that occurs in multicellular organisms. It is well established that dysregulation of apoptosis plays an important role in pathogenesis of malignant diseases and may be associated with tumor progression and resistance to cytotoxic treatment. This study aimed to evaluate expression of apoptosis inducing factor (AIF) in GCTs. We observed lower AIF expression in GCTs compared to normal testicular tissue. We also showed prognostic significance of AIF in GCTs. AIF downregulation might represent one of the mechanisms of inhibition of apoptosis with subsequent facilitation of cell survival and metastatic dissemination of GCTs and perhaps could serve as a potential therapeutic target. Apoptosis is a strictly regulated process essential for preservation of tissue homeostasis. This study aimed to evaluate expression of apoptosis inducing factor (AIF) in testicular germ cell tumors (GCTs) and to correlate expression patterns with clinicopathological variables. Formalin-fixed and paraffin-embedded specimens of non-neoplastic testicular tissue and GCTs obtained from 216 patients were included in the study. AIF expression was detected by immunohistochemistry, scored by the multiplicative quickscore method (QS). Normal testicular tissue exhibits higher cytoplasmic granular expression of AIF compared to GCTs (mean QS = 12.77 vs. 4.80, p < 0.0001). Among invasive GCTs, mean QS was the highest in embryonal carcinoma, yolk sac tumor and seminoma, lower in teratoma and the lowest in choriocarcinoma. No nuclear translocation of AIF was observed. Nonpulmonary visceral metastases were associated with lower AIF expression. Metastatic GCTs patients with high AIF expression had better overall survival compared to patients with low AIF expression (HR = 0.26, 95% CI 0.11–0.62, p = 0.048). We observed significantly lower AIF expression in GCTs compared to normal testicular tissue, which is an uncommon finding in malignant tumors. AIF downregulation might represent one of the mechanisms of inhibition of apoptosis and promotion of cell survival in GCTs.

Commentary to ‘Oncologic outcomes of pre-malignant and invasive germ cell tumors in patients with differences in sex development – A systematic review’

Commentary to ‘Oncologic outcomes of pre-malignant and invasive germ cell tumors in patients with differences in sex development – A systematic review’

Oncologic outcomes of pre-malignant and invasive germ cell tumors in patients with differences in sex development – A systematic review

To describe the rates of GCNIS-free and GCT-free pathology based on age at gonadal surgery and to describe long-term oncologic outcomes in patients with DSD who have GCNIS or GCT at the time of gonadal surgery. A systematic review was conducted using MEDLINE to identify patients with DSD who underwent gonadal surgery. DSD diagnoses were stratified based on malignancy risk. GCNIS/GCT and GCT-free survival by age of gonadal surgery, RFS and OS were calculated using the Kaplan-Meier method, with groups compared using log-rank testing. 386 articles from 1951 to 2017 were included (2037 patients). Median age at gonadal surgery was 17 years (y) (IQR 11-20), median follow-up was 60 months (m) (IQR 30-68.1). GCNIS/GCT- and GCT-free survival at the time of gonadal surgery was lowest for those in the high/intermediate risk group (p<0.001) but decreased sharply around age 15y, regardless of risk category. 5y RFS and OS was similar for those with no GCNIS/GCT and GCNIS and was worse for those with GCT (p<0.001). When patients undergo gonadal surgery, regardless of indication (i.e. prophylactic vs. tumor), it appears that GCTs are more commonly found when surgery is done around age 15y or older, despite risk category. This is similar to ovarian and testicular GCTs. Patients with GCNIS can be reassured that long-term oncologic outcomes are excellent. While RFS and OS for GCTs are not as good as for ovarian and testicular GCTs (95%), they are still >80%. This similar trend was found in a COG review of 9 patients with DSD and ovarian GCT. There were several limitations to this study. This is a retrospective analysis that included aa wide time frame of publications. The indication for surgical intervention was not addressed in the majority of publications. Thus these data provide pathologic outcomes based on age at gonadal surgery rather than the age at which GCNIS/GCT develops over a lifetime, if at all. The risk of GCNIS or GCT at the time of gonadal surgery appears to increase with age, accelerating between 15 and 20y regardless of risk category. 5y RFS and OS for those with GCNIS is equivalent to those without GCNIS/GCT but is worse for those with GCT. These data may be used when counseling families on timing of gonadal surgery and quantification of outcomes should GCNIS or malignancy be identified.

Testicular epidermoid cysts: a reevaluation

BackgroundTesticular epidermoid cysts (TECs) are rare benign testicular neoplasms. As TECs are rarely associated with germ cell tumours (GCTs), the understanding of biological behaviour and clinical management of TEC is unresolved.MethodsWe retrospectively searched the files of patients treated for testicular neoplasms and germ cell cancer in the time from 2000 to 2017. Those with TEC were subjected to closer review looking to clinical and histological features, and to results from imaging with ultrasonography (US), contrast enhanced sonography (CEUS) and magnetic resonance imaging (MRI).ResultsAmong 589 patients undergoing surgery for testicular tumour, nine simple TECs were identified (1.5, 95% confidence intervals 0.53–2.50%). Median age was 26 years. Imaging revealed sharply demarcated roundish lesions with avascular central areas. Eight patients underwent testis-sparing excision with no recurrence ensuing. One had orchiectomy because of large size of the mass. Histologically, TECs consisted of cornifying squamous cell epithelium and no accompanying germ cell neoplasia in situ. Two additional cases (0.3% of all) required orchiectomy because these TECs were associated with ipsilateral GCT.ConclusionsTEC is usually a benign lesion that can safely be diagnosed with US, CEUS and MRI due to its roundish shape and its avascular centre. Histologically, this TEC corresponds to the prepubertal-type teratoma unrelated to germ cell neoplasia in situ of the 2016 WHO classification. The other subtype of TEC that is associated with invasive GCT represents a teratoma of postpubertal-type. From a clinical point of view it could be easier to differentiate between a “simple TEC” which is benign (prepubertal type) and a “complex TEC” which is malignant because of its association with invasive GCT.

Immunolocalization of DMRTB1 in human testis with normal and impaired spermatogenesis.

The transcription factor DMRTB1 plays a pivotal role in coordinating the transition between mitosis and meiosis in murine germ cells. No reliable data are available for human testis. The present study aims to examine the testicular expression pattern of DMRTB1 in men showing normal and impaired spermatogenesis. Immunohistochemistry was performed using 54 human testicular biopsy specimens and a commercial rabbit polyclonal anti-DMRTB1 primary antibody. RT-PCR complemented immunohistochemistry. To further characterize immunopositive cells and possible co-localization, the proliferation marker Ki-67, the tumor marker PLAP, and an anti-DMRT1 antibody were used. In men with normal spermatogenesis, a strong immunoreactivity was detectable in a subset of spermatogonia (38.34±2.14%). Some spermatocytes showed a weak immunostaining. Adjacent Sertoli cells were immunonegative. Compared with a hematoxylin and eosin overview staining, these immunopositive cells were almost exclusively identified as Apale and B spermatogonia and primary spermatocytes in (pre-)leptotene, zygotene, and pachytene stages. In patients with spermatogenic arrest at spermatogonial level, an altered staining pattern was found. No immunoreactivity was detected in Sertoli cells in Sertoli cell-only syndrome. In germ cell neoplasia insitu (GCNIS) tubules, except for a few (0.4±0.03%), pre-invasive tumor cells were immunonegative. Seminoma cells showed no immunostaining. According to previous findings in mice, it seems reasonable that DMRTB1 is expressed in these normal germ cell populations. Moreover, altered staining pattern in spermatogenic arrest at spermatogonial stage suggests a correlation with mitosis and transformation into B spermatogonia. The absence of DMRTB1 in GCNIS cells and tumor cells might be associated with uncontrolled neoplastic cell proliferation and progression into invasive germ cell tumors. Further research is required to elucidate, for example, the role of DMRTB1 in the malignant transformation of human germ cells. Our data indicate a relevant role for DMRTB1 regarding the entry of spermatogonia into meiosis in men.

Gonadoblastoma in patients with 45,X/46,XY mosaicism: A 16-year experience

It is recognised that individuals with a 45,X/46,XY karyotype, known as Turner mosaic syndrome with Y chromosome material (TMSY), have an increased risk of developing gonadoblastoma (GB), which may then devolve into one of a number of germ cell malignancies. Hence, children with TMSY are usually recommended to undergo prophylactic gonadectomy. We designed this study to describe the phenotypic features of our series of children with TMSY who underwent prophylactic gonadectomy in order to evaluate the prevalence of GB and germ cell malignancies in their resected specimens. This is a retrospective case series wherein we comprehensively reviewed the clinical, histological, and cytogenetic features of all patients who underwent prophylactic gonadectomy at three tertiary paediatric referral centres over 16 years. Cases were identified from surgical logbooks and through the institutional histopathology database. Data were collected with particular reference to clinical phenotype, predominant karyotype cell line, operative management, anatomical findings and the presence of neoplastic changes. Fourteen children ranging in age at the time of surgery from 2 weeks to 17 years were included in the series. Eleven children were reared as females. The three children who were reared as males had severe penoscrotal hypospadias. The 46,XY cell line was the predominant cell line in seven (50%) cases in blood lymphocytes. The resected specimens from four patients (28.6%) contained GB, with three patients having bilateral GB. This sub-group of patients with GB were aged 5 months, 48 months, 71 months, and 13 years. GB arose in one patient with and three patients without genital virilisation. There was no focus of invasive germ cell tumour in any specimen. GB may be present in infants with TMSY as young as 5 months, even with low levels of Y chromosome material. The prevalence of GB in prophylactic gonadectomy specimens is similar to many previously reported series, although the absence of dysgerminoma in our series is reassuring. The exclusive presence of GB in intra-abdominal gonads is in keeping with the findings of several other series. Owing to the presence of gonadoblastoma in the gonads of children with TMSY as young as 5 months, we recommend that all patients with intra-abdominal gonads in the context of TMSY should duly undergo prophylactic gonadectomy, although the timing of such surgery can be discussed with parents during counselling regarding the risk of malignancy.

The risk of neoplasm associated with dysgenetic testes in prepubertal and pubertal/adult patients.

In patients with Y-chromosome in the karyotype, partial gonadal dysgenesis and disorders of male reproductive sex organs development are usually resected in childhood because of the high risk of germ cell tumours (GCT). In patients with Y-chromosome, complete gonadal dysgenesis and female genitalia gonadectomy is performed markedly later. However, due to the relatively low number of adult patients with preserved dysgenetic gonads, the true risk of neoplasm is unknown. The aim of the study was to evaluate the prevalence of neoplasia in dysgenetic gonads of children and adults with Y-chromosome in a retrospective study. A review of medical documentation of 94 patients with disorders of sex development (DSD), Y-chromosome and gonadal dysgenesis (GD), aged 1.2-32 years (47 prepubertal, 1.2-10 years; 47 pubertal/adult, 13-32 years), was conducted. Serum levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH) and testosterone were determined. Bilateral gonadectomy was performed in 73.4% of patients, and unilateral gonadectomy with biopsy of the contralateral gonad in 26.4%. All gonadal tissues were subjected to immunohistochemical evaluation with antibodies against PLAP and OCT3/4 (markers of malignant germ cells, but also foetal multipotent germ cells), while gonads of prepubertal patients were examined by c-KIT, as well. Streak gonads were identified on both sides (complete GD) in 30.8%, a streak gonad on one side and an underdeveloped testis on the other (asymmetric GD) in 38.3%, and underdeveloped testicular structure on both sides (partial GD) in 30.8% of cases. Germ cell neoplasia was found in 53.2% of patients (51.1% in children, 55.3% in pubertal/adults). Invasive GCT were identified in 11.7% of cases, of which 90.9% were in pubertal/adult patients. Other neoplastic lesions included gonadoblastoma (16% prevalence) and testicular carcinoma in situ (25.5%). In younger patients FSH serum levels were increased in 81% of cases (mean 2.82 ± 2.18 IU/L), while LH in 58% (mean 1.82 ± 1.69 IU/L). Hypergonadotropic hypogonadism was diagnosed in most of the pubertal/ /adult patients (mean FSH 54.2 ± 23.3 IU/L, mean LH 21.7 ± 12.1 IU/L, mean testosterone 5.5 ± 4.5 nmol/L). Dysgenetic gonads in patients with Y chromosome have a high risk of germ cell neoplasia (ca. 50%). If they are preserved until puberty/early adulthood, they may develop overt, invasive GCT. The gonads also have poor hormonal activity (hypergonadotropic hypogonadism) in most of the pubertal/adult patients. Each of these cases must be considered individually and a decision to remove the gonad or not should be based on the comprehensive analysis of the phenotype by a multidisciplinary team of specialists in consultation with the patient and the parents. If dysgenetic gonads are not resected in childhood, these patients need careful ongoing follow-up examination, including biopsy and histopathological evaluation.

Gonadoblastoma and hepatoid and endometrioid-like yolk sac tumor: an update.

Dr Robert E. Scully greatly advanced our understanding of germ cell neoplasia to the extent that it is difficult to narrow the discussion of his contributions to this topic so that it can be covered in a brief article. This article accordingly focuses on some of the recent developments concerning 2 of his major contributions in this area-the gonadoblastoma (GB) and variant morphologies of yolk sac tumor. GB was defined by Dr Scully in 1953 and its features elaborated in detail by him in 1970. This neoplasm occurred in young patients who often displayed phenotypic sex ambiguities and frequently presented with primary amenorrhea. It was bilateral in 40%, and consisted of circumscribed nests of small sex cord cells and germinoma-like cells admixed with round deposits of eosinophilic, hyaline, often calcified material. These nests were set in a spindle cell gonadal stroma with Leydig-like or lutein-like cells. Because of his work we now understand that this precursor to invasive germ cell tumors occurs in patients with a specific form of disorder of sex development, namely gonadal dysgenesis, and only in those who have a particular portion of the Y chromosome, the GB locus/TSPY gene, within the gonadal tissue. An essential element to the development of GB appears to be a defect in the genetic pathway that leads to the development of Sertoli cells. Improperly formed Sertoli cells predispose to "delayed maturation" of the gonocytes of the gonad and predispose them to undergo malignant transformation. "Undifferentiated gonadal tissue" has been proposed as the precursor to the development of GB and consists of an unorganized mixture of apparently non-neoplastic germ cells, germ cells with delayed maturation, and neoplastic germ cells with sex cord cells and gonadal stroma. Two variant morphologies of yolk sac tumor were also recognized by Dr Scully. In the hepatoid variant features similar to hepatocellular carcinoma occurred, although primitive glandular foci and lack of liver involvement permitted its distinction in most cases. More recently this variant has been found to occasionally produce bile in canalicular-like structures and to stain strongly for both SALL4 and glypican 3, 2 recently described markers of yolk sac tumor. Recognition of hepatoid yolk sac tumor was followed by the description of a potential mimic, primary ovarian hepatoid carcinoma, which, however, occurred in a significantly older patient population and was occasionally associated with surface epithelial neoplasia. The endometrioid-like variant of yolk sac tumor simulated primary endometrioid adenocarcinoma. It can be suspected on routine stains because of primitive appearing nuclei, frequent subnuclear vacuoles, and in some cases association with more usual yolk sac tumor. Its recognition is now facilitated by a panel of immunohistochemical stains that are often expressed differentially in these 2 neoplasms--endometrioid-like yolk sac tumor: positive for SALL4, glypican 3, and α-fetoprotein; endometrioid adenocarcinoma: positive for cytokeratin 7 and epithelial membrane antigen. Finally, Dr Scully contributed one of the first cases in the literature of yet another nuance in the complicated world of yolk sac neoplasia, namely the development of some tumors on the background of a surface epithelial neoplasm. This is analogous to the more common development of choriocarcinoma from carcinoma and, in the case of yolk sac tumor, diagnosis is aided clinically by the usual older age of the patient and nature of the associated neoplasia.

Expression of SALL4 and SF-1 in Gonadoblastoma

Gonadoblastoma is a rare gonadal neoplasm composed of primordial germ cells and sex cord-stromal cells and usually occurs in patients with dysgenetic gonads. When patients with gonadoblastoma develop an invasive germ cell tumor, the invasive germ cell component can take the form of dysgerminoma/seminoma, embryonal carcinoma, or yolk sac tumor. In this study, we performed immunohistochemical analysis for SALL4 and steroidogenic factor-1 (SF-1) on 4 cases of gonadoblastoma to examine the expression patterns of these proteins. All of the patients were phenotypically female. One patient had Swyer syndrome, the rest had Turner syndrome. The primordial germ cell component was histologically similar to cells in dysgerminoma/seminoma in these 4 cases. Two patients showed the invasive component-dysgerminoma. As expected, SALL4 stained the germ cells and SF-1 stained the sex cord-stromal cells. There was a clear distinction between the staining patterns of these 2 cell populations. This study demonstrates the utility of SALL4 and SF-1 in determining whether or not there is an invasion in the primordial germ cell component.

The Transdifferentiation of Mediastinal Germ Cell Tumor into a Myeloid Neoplasm in the Bone Marrow-Report of a Case and Short Review of a Diagnostic Pitfall

We present the case of a young male with a mediastinal non-seminomatous germ cell tumor who developed a severe thrombocytopenia, one year after initial diagnosis and surgical treatment and chemotherapy. Bone marrow morphology suggested myelodysplastic syndrome (refractory cytopenia with multilineage dysplasia). Cytogenetics of bone marrow showed near tetraploid karyotype, raising concerns about the diagnosis of MDS. The patient died within one month due to refractory disease and autopsy revealed a highly invasive germ cell tumor in the mediastinum and chest wall which showed the same cytogenetic abnormality seen in the bone marrow. The bone marrow collected at autopsy showed features compatible with acute myeloid leukemia. The implications of this transdifferentiation of a mediastinal germ cell tumor into an apparent hematological malignancy for correct diagnosis and appropriate treatment are discussed.

The downside of ‘inappropriate messaging’: new insight into the development of testicular germ cell tumours in young men?

How invasive testicular germ cell tumours (TGCTs) develop from precursor carcinoma in situ/intratubular germ cell neoplasia unclassified (CIS/IGCNU) cells, and only after puberty, is unknown. In the current issue of The Journal of Pathology, Jørgensen and colleagues have compared the protein expression profile of CIS before and after puberty and in pre-invasive versus invasive TGCT and show that the mitosis-meiosis controller DMRT1 switches off in CIS cells postpubertally and is associated with invasiveness. They also show that CIS cells express a 'confusing' mix of pro- and anti-meiotic proteins; this may predispose CIS cells to accumulate extra chromosomal material which ultimately leads to tumourigenesis.

Phase II study of everolimus (E) in refractory germ cell tumors (GCTs).

TPS4677 Background: GCTs represent a model for the cure of cancer. Nonetheless, a small proportion of patients (pts) develop disease recurrence. Because of insufficient results in the treatment of relapsed GCTs, evaluation of new treatments is a priority. Loss of the tumor suppressor gene PTEN (phosphatase and tensin homolog) marks the transition from intratubular germ cell neoplasias (ITGCN) to invasive GCTs. In the testis, PTEN was abundantly expressed in germ cells whereas it was virtually absent from 56% of seminomas as well as from 86% of embryonal carcinomas and virtually all teratomas. On the contrary, ITGCN intensely expressed PTEN, indicating that loss of PTEN expression is not in early event in GCTs development, but is associated with disease progression. PTEN inactivation is associated with dysregulation of PI3K/Akt pathway and increased mTOR signalling. We hypothesize that dysregulation of PI3K/Akt pathway due to PTEN inactivation in GCTs suggests that these pts could have greater benefit from mTOR inhibition. Methods: In December 2011, National Cancer Institute of Slovakia launched an one arm, two-staged phase II study aimed to evaluate the efficacy and toxicity of E in pts with refractory GCTs. The primary objective is to determine the efficacy of E in patients with refractory GCTs. Secondary objectives includes favourable response rate, time to progression and safety. The pts with radiological and/or serological proof of relapsed GCTs, who were not amenable to be cured by chemotherapy or surgery and who failed at least two platinum-based regimens or one platinum regimen in case of platinum-refractory disease or primary mediastinal non-seminomatous GCTs were eligible. All other standard inclusion and exclusion criteria for study of salvage treatment in refractory GCTs pts were applied. E will be administered at a dose of 10mg daily until progression or unacceptable toxicity. Assuming a response rate of clinical interest of ≥40%, a minimal response rate of 20%, a probability of 5% for rejecting an active drug, and a probability of 20% to further evaluate an ineffective drug, 18 pts will be enrolled into the first cohort. If &lt;4 responses will be observed, the study will be terminated, otherwise, it will be continued with a second cohort of 20 pts.

A genome-wide RNAi screen in mouse embryonic stem cells identifies Mp1 as a key mediator of differentiation

Despite intense investigation of intrinsic and extrinsic factors that regulate pluripotency, the process of initial fate commitment of embryonic stem (ES) cells is still poorly understood. We used a genome-wide short hairpin RNA screen in mouse ES cells to identify genes that are essential for initiation of differentiation. Knockdown of the scaffolding protein Mek binding protein 1 (Mp1, also known as Lamtor3 or Map2k1ip1) stimulated self-renewal of ES cells, blocked differentiation, and promoted proliferation. Fibroblast growth factor 4 (FGF4) signaling is required for initial fate commitment of ES cells. Knockdown of Mp1 inhibited FGF4-induced differentiation but did not alter FGF4-driven proliferation. This uncoupling of differentiation and proliferation was also observed when oncogenic Ras isoforms were overexpressed in ES cells. Knockdown of Mp1 redirected FGF4 signaling from differentiation toward pluripotency and up-regulated the pluripotency-related genes Esrrb, Rex1, Tcl1, and Sox2. We also found that human germ cell tumors (GCTs) express low amounts of Mp1 in the invasive embryonic carcinoma and seminoma histologies and higher amounts of Mp1 in the noninvasive carcinoma in situ precursor and differentiated components. Knockdown of Mp1 in invasive GCT cells resulted in resistance to differentiation, thereby showing a functional role for Mp1 both in normal differentiation of ES cells and in germ cell cancer.

Embryonic Stem Cell Transplantation into Seminiferous Tubules: A Model for the Study of Invasive Germ Cell Tumors of the Testis

Over the last 15 years, cell transplantation into seminiferous tubules has become a valuable tool to study germinal cell biology and related matters. This is particularly so, because the blood-testis permeability barrier establishes a sealed compartment which protect against certain influences such as immunological rejection. In the light of the functional and genetic similarities between carcinoma in situ (CIS) of the testis and embryonic stem (ES) cells, our laboratory has developed a tumor assay to study cancer invasion processes in testicular germ cell tumors (TGCT) based on the transplantation of ES cells into the seminiferous tubules. Here, we describe this new tumor assay and provide additional information regarding the transplantation techniques used and their application for the study of TGCTs. Finally, we discuss the practical implications of our experimental approach and its potential application for the understanding of TGCT invasive processes and the development of new antineoplastic strategies.

Diagnosis of testicular carcinoma in situ‘(intratubular and microinvasive)’ seminoma and embryonal carcinoma using direct enzymatic alkaline phosphatase reactivity on frozen histological sections

Testis-sparing surgery might benefit quality of life, but can only be applied with histological examination for the presence of invasive germ cell tumour components, and the precursor carcinoma in situ (CIS). Currently, diagnosis is based on paraffin-embedded tissue, therefore a delay in further surgery is mainly unavoidable. The aim was to develop an intraoperative assessment technique using alkaline phosphatase as a marker. A total of 4093 snap-frozen samples and matched paraffin-embedded tissue of 1500 patients were included. Besides standard haematoxylin and eosin (H&E) staining, the direct enzymatic alkaline phosphatase reactivity (dAP) test (duration 15 min) was applied on frozen sections, while H&E and immunohistochemistry for detection of OCT3/4, α-fetoprotein, human chorionic gonadotrophin (hCG) and cytokeratin was performed on the paraffin-embedded slides. Endothelial cells served as control for the dAP test. Positive staining was found in all CIS (n=965), seminoma (n=1035) and embryonal carcinoma (n=584), either intratubular, microinvasive or invasive. Differentiated non-seminomas (n=1238) showed variable staining. No staining was identified in spermatocytic seminomas (n=5), testicular lymphomas (n=42), testicular rhabdomyosarcomas (n=7), Leydig cell tumours (n=31), Sertoli-cell-only nodules (n=4), (epi) dermoid cyst (n=16), normal testicular parenchyma (n=116), testicular torsion (n=32) and inflammation of the epididymis (n=19). The dAP test results matched H&E-stained parallel sections, as well paraffin-embedded tissue analysis, including immunohistochemistry. The dAP test is an informative, reproducible and easy tool to diagnose CIS, (intratubular and microinvasive) seminoma and embryonal carcinoma on frozen tissue sections, being of great value in the context of sparing surgery.

Contralateral testicular biopsy in testis cancer: current concepts and controversies

Of all patients with unilateral testis cancer, approximately 5% harbour testicular intraepithelial neoplasia (TIN) in their contralateral testicle that will progress into an invasive germ cell tumour over time. The accurate diagnosis of TIN by a random two-site surgical testis biopsy and effective therapy by local radiation has led to the concept of a contralateral screening biopsy in all patients with testis cancer. However, screening and preventive treatment are only indicated if the therapeutic outcome of the screened population is improved, and the physiological function of the affected organ is not impaired. Based on a critical review of previous reports, some drawbacks of this policy have to be considered and question the routine indication for contralateral testis biopsy: (i) all TIN-negative patients still have to undergo meticulous follow-up for metachronous testis cancer due to a false negative biopsy rate of 0.5-1.0%; (ii) local radiation of TIN results in irreversible infertility due to eradication of spermatogenesis; (iii) local radiation of TIN results in an impairment of endocrine Leydig cell function in 25% of the patients; (iv) therapeutic outcome and prognosis will not be improved in irradiated patients as compared to patients on surveillance; (v) local tumour resection for the management of metachronous testicular cancer represents an effective and viable option. Current reports do not support the strategy of contralateral testis biopsy in all patients with unilateral testicular germ cell tumours. According to the recommendations of the European Germ Cell Cancer Consensus Group, a testis biopsy might be offered to high-risk patients for contralateral TIN (testicular volume <12 mL, history of cryptorchidism, age <30 years).

Analysis of Gene Expression Profiles of Microdissected Cell Populations Indicates that Testicular Carcinoma In situ Is an Arrested Gonocyte

Testicular germ cell cancers in young adult men derive from a precursor lesion called carcinoma in situ (CIS) of the testis. CIS cells were suggested to arise from primordial germ cells or gonocytes. However, direct studies on purified samples of CIS cells are lacking. To overcome this problem, we performed laser microdissection of CIS cells. Highly enriched cell populations were obtained and subjected to gene expression analysis. The expression profile of CIS cells was compared with microdissected gonocytes, oogonia, and cultured embryonic stem cells with and without genomic aberrations. Three samples of each tissue type were used for the analyses. Unique expression patterns for these developmentally very related cell types revealed that CIS cells were very similar to gonocytes because only five genes distinguished these two cell types. We did not find indications that CIS was derived from a meiotic cell, and the similarity to embryonic stem cells was modest compared with gonocytes. Thus, we provide new evidence that the molecular phenotype of CIS cells is similar to that of gonocytes. Our data are in line with the idea that CIS cells may be gonocytes that survived in the postnatal testis. We speculate that disturbed development of somatic cells in the fetal testis may play a role in allowing undifferentiated cells to survive in the postnatal testes. The further development of CIS into invasive germ cell tumors may depend on signals from their postpubertal niche of somatic cells, including hormones and growth factors from Leydig and Sertoli cells.

Multifocality in Testicular Germ Cell Tumors

Standard treatment for testicular germ cell tumor is radical orchiectomy. Several groups have suggested an organ sparing approach in patients with bilateral tumors or tumor in a solitary testis. We determined the prevalence of multifocality in testicular germ cell tumor cases. Orchiectomy specimens from 145 consecutive patients treated for testicular germ cell tumor between 1995 and 2006 were included in the study. In the current series slides were reviewed by a single dedicated uropathologist. Multifocality was defined as 1 of 4 distinct pathological entities, including 1) distinct tumor focus conspicuously separable from the main tumor mass, 2) microinvasive tumor characterized by a single or small groups of malignant germ cells scattered within the normal interstitial parenchyma, 3) extra tumor vascular invasion and 4) rete testis invasion by pagetoid tumor spread. Multifocality was identified in 48 patients (33%), of whom 17 (12%) had an additional distinct tumor focus, 21 (14%) had microinvasive tumor, 17 had extra tumor vascular invasion and 2 had rete testis invasion by pagetoid tumor spread. Multifocality was more prevalent in men with smaller tumors and seminomatous histology (pure seminoma or as part of a mixed germ cell tumor). Multifocality was present in 63% of men with an index mass of 2 cm or less in diameter. Study limitations include potential pathological sampling errors resulting from the retrospective design. Multifocality is a frequent finding in testicular germ cell tumor cases that is associated with small mass size and seminomatous histology. Data suggest that additional invasive tumor outside the index mass may be present in up to 63% of men considered potential candidates for organ sparing surgery.

Contralateral intratubular germ cell neoplasia in a patient with testicular cancer

A 27-year-old man was referred to an oncology department following right orchiectomy for a stage I testicular seminoma at high risk for recurrence. He presented 6 weeks after the orchiectomy with an atrophic left testis, fatigue and a history of infertility. Measurement of serum levels of urea, electrolytes, liver enzymes, bilirubin, human chorionic gonadotropin, alpha-fetoprotein, lactate dehydrogenase, testosterone and luteinizing hormone, full blood count, and left testicular biopsy. Tubular atrophy of the left testis with islands of intratubular germ cell neoplasia (ITGCN), and hypergonadotropic hypogonadism. The patient received adjuvant chemotherapy as a single dose of carboplatin for the seminoma at high risk for recurrence, and testosterone replacement for the hypergonadotropic hypogonadism. Radiotherapy to the ITGCN-bearing solitary testis or a second orchiectomy was offered to prevent the progression of ITGCN into an invasive germ cell tumor. After exploring his options with regards to fertility treatment, the patient chose to undergo second orchiectomy with a subsequent, unsuccessful, attempt at sperm retrieval. At 20 months after diagnosis of his initial seminoma the patient showed no sign of recurrence.